A double sandwich monoclonal enzyme immunoassay for detection of hepatitis B surface antigen.

An enzyme immunoassay for detection of hepatitis B surface antigen based on the use of 3 monoclonal antibodies (mAbs) was developed: an IgM as capture and 2 IgG1 for detection. The system biotin-streptavidin was compared with direct conjugation of mAbs to peroxidase and was preferred because of its higher signal to noise ratio. The possibility of simultaneous addition of human serum and biotin-mAb was discarded because of an evident prozone effect with some sera containing high HBsAg levels. The conjugation of biotin to IgG1 mAbs through a spacer arm (amidocaproyl) and the use of a highly sensitive substrate (tetramethylbenzIdine) improved the assay detection limit by about 10 times.